1. Field of the Invention
The present invention relates to an in vitro testing method for studying the penetration behavior of dermal products. More particularly, the present invention relates to the use of a specific model barrier membrane in the study of percutaneous penetration of bioactive agents.
2. Description of the Prior Art
The primary function of the skin of a mammalian organism, including humans, is to protect the body from foreign substances and to control the loss of water, electrolytes and other body constituents. Human skin is mainly composed of three layers. The first layer from the outer surface is the stratum corneum, then the living epidermis and the dermis. In comparison with the last two layers, the stratum corneum has been found to be much less permeable to relatively polar molecules and acts as the primary barrier against the transfer of such molecules across the skin.
The stratum corneum is composed of a mass of dead epidermal cells which in turn consists mainly of keratinized proteins and lipids as well as moisture. Alternate intracellular and intercellular layers comprise the multilamellar structure of the stratum corneum. Its total thickness can vary widely, but averages approximately 10-20 microns. The origin of the intracellular portion is the endogenous material of the living epidermal cell containing mainly keratinized protein. On the other hand, the intercellular portion contains lipids, principally triglycerides, fatty acids, cholesterol and phospholipid, which are derived from epidermal lamellar bodies and nonfibrous protein. Recent work has shown that the barrier function against water molecules is apparently located primarily in the intracellular portion of the stratum corneum.
Due to the barrier properties of the skin, in the development of pharmaceutical products which are intended to be applied to the skin and which provide medications for delivery into the skin, it is highly desirable to have available an in vitro testing method to study the penetration behavior of such dermal products. Indeed, in vitro testing of the availability of bioactive agents from formulations applied to the skin is widely practiced. Due to the lack of a suitable model barrier membrane, however, conventional testing methods have not proven highly satisfactory.
In general, human skin has often been used in in vitro studies since it is obviously the best model barrier membrane to predict the effectiveness of transdermal absorption. Human cadaver skin has generally been used in such instances. Certain animal skins whose properties are thought to be similar to human skin have also been used in many in vitro percutaneous transport studies. The skins of relatively hairless mammals such as the fuzzy rat, the hairless mouse and the baby pig, for example, have been used. The major problem encountered with using cadaver skin or fresh animal skin, however, is the great variability encountered with different samples. Human skin shows particularly large variations in its permeability depending upon site, age, etc. Membrane skin specimens from rats and mice also show similar variations.
Another problem with cadaver skin membrane and fresh skin samples from animals is that they undergo rapid deterioration, sometimes over the period of the test itself, which may last several days. Such problems with decomposition require that the cadaver skin be used within forty-eight hours. Another serious problem is that various substances are released from such fresh tissues, e.g., lipids, which substances interfere with direct analysis of the chemical substance being followed, e.g., by UV spectrophotometric analysis.
Artificial membranes have also been used for in vitro penetration studies because of their uniformity and hydrophobic character, which resembles the stratum corneum of human skin. Their suitability has been found to be less than satisfactory, however, particularly when used with formulations containing dermal enhancers.
Thus, there is a need for a model barrier membrane which closely matches the permeability of the human stratum corneum, but which is also readily available and provides uniform results. The use of such a membrane in in vitro transdermal absorption testing would greatly enhance the test results and ones ability to run the tests.
Accordingly, there is provided by the present invention a novel method for in vitro testing the ability of formulations to deliver bioactive agents across skin.
Another object of the present invention is to provide such a test method which employs a novel barrier or model membrane.
Still another object of the present invention is to provide such a test method which employs a model test membrane comprised of a material which has a permeability profile which closely matches the permeability of the human stratum corneum.
Yet another object of the present invention is to provide such a novel test method which uses a model test membrane comprised of a material which is readily available and provides uniform results.
Another object of the present invention is to provide such a test method which uses a model test membrane comprised of a material which can be stored for extended periods of time.
These and other objects, as well as the scope, nature and utilization of the invention, will be apparent to those skilled in the art from the following description and the appended claims.